@article{walsh_tollison_brochu_shaw_diveley_chou_law_kirk_gale_peng_2022, title={Single-Cell-Based High-Throughput Ig and TCR Repertoire Sequencing Analysis in Rhesus Macaques}, volume={208}, ISSN={["1550-6606"]}, url={https://doi.org/10.4049/jimmunol.2100824}, DOI={10.4049/jimmunol.2100824}, abstractNote={Abstract
               Recent advancements in microfluidics and high-throughput sequencing technologies have enabled recovery of paired H and L chains of Igs and VDJ and VJ chains of TCRs from thousands of single cells simultaneously in humans and mice. Despite rhesus macaques being one of the most well-studied model organisms for the human adaptive immune response, high-throughput single-cell immune repertoire sequencing assays are not yet available due to the complexity of these polyclonal receptors. We used custom primers that capture all known rhesus macaque Ig and TCR isotypes and chains that are fully compatible with a commercial solution for single-cell immune repertoire profiling. Using these rhesus-specific assays, we sequenced Ig and TCR repertoires in >60,000 cells from cryopreserved rhesus PBMCs, splenocytes, and FACS-sorted B and T cells. We were able to recover every Ig isotype and TCR chain, measure clonal expansion in proliferating T cells, and pair Ig and TCR repertoires with gene expression profiles of the same single cells. Our results establish the ability to perform high-throughput immune repertoire analysis in rhesus macaques at the single-cell level.}, number={3}, journal={JOURNAL OF IMMUNOLOGY}, author={Walsh, Evan S. and Tollison, Tammy S. and Brochu, Hayden N. and Shaw, Brian I and Diveley, Kayleigh R. and Chou, Hsuan and Law, Lynn and Kirk, Allan D. and Gale, Michael, Jr. and Peng, Xinxia}, year={2022}, month={Feb}, pages={762–771} }
 @article{brochu_tseng_smith_thomas_jones_diveley_law_hansen_picker_gale_et al._2020, title={Systematic Profiling of Full-Length Ig and TCR Repertoire Diversity in Rhesus Macaque through Long Read Transcriptome Sequencing}, volume={204}, ISSN={["1550-6606"]}, DOI={10.4049/jimmunol.1901256}, abstractNote={Abstract
               The diversity of Ig and TCR repertoires is a focal point of immunological studies. Rhesus macaques (Macaca mulatta) are key for modeling human immune responses, placing critical importance on the accurate annotation and quantification of their Ig and TCR repertoires. However, because of incomplete reference resources, the coverage and accuracy of the traditional targeted amplification strategies for profiling rhesus Ig and TCR repertoires are largely unknown. In this study, using long read sequencing, we sequenced four Indian-origin rhesus macaque tissues and obtained high-quality, full-length sequences for over 6000 unique Ig and TCR transcripts, without the need for sequence assembly. We constructed, to our knowledge, the first complete reference set for the constant regions of all known isotypes and chain types of rhesus Ig and TCR repertoires. We show that sequence diversity exists across the entire variable regions of rhesus Ig and TCR transcripts. Consequently, existing strategies using targeted amplification of rearranged variable regions comprised of V(D)J gene segments miss a significant fraction (27–53% and 42–49%) of rhesus Ig/TCR diversity. To overcome these limitations, we designed new rhesus-specific assays that remove the need for primers conventionally targeting variable regions and allow single cell level Ig and TCR repertoire analysis. Our improved approach will enable future studies to fully capture rhesus Ig and TCR repertoire diversity and is applicable for improving annotations in any model organism.}, number={12}, journal={JOURNAL OF IMMUNOLOGY}, author={Brochu, Hayden N. and Tseng, Elizabeth and Smith, Elise and Thomas, Matthew J. and Jones, Aiden M. and Diveley, Kayleigh R. and Law, Lynn and Hansen, Scott G. and Picker, Louis J. and Gale, Michael, Jr. and et al.}, year={2020}, month={Jun}, pages={3434–3444} }